Electron microscopic structure and innervation of the carotid baroreceptor region in the rock hyrax (Procavia capensis).

Semi-thin plastic sections reveal that the carotid baroreceptor region in the rock hyrax comprising the origin of the internal carotid artery has a preponderantly elastic structure and a thick tunica adventitia. In contrast, the common carotid artery has a musculoelastic structure, whereas the cranial segment of the internal carotid artery (immediately distal to the baroreceptor areas) shows the features of a muscular artery. Electron microscopy discloses the presence of sensory nerve endings within the parts of the tunica adventitia adjoining the preponderantly elastic zone of the internal carotid artery. These nerve endings are characterized by varicose regions containing a large quantity of mitochondria. Bundles of collagen fibers in the tunica adventitia form convolutions or whorls around the nerve terminals and often terminate on the surface of the elastic fibers or into the basement membranes of the neuronal profiles. The large content of elastic tissue in the tunica media of the baroreceptor region may render the vessel wall highly distensible to intraluminal pressure changes. This, in turn, would facilitate the transmission of the stimulus intensity to the sensory nerve terminals located in the tunica adventitia. It is suggested that the stretching of elastic fibers may form the main mechanical event leading to the distortion of the associated nerve terminals. However, a change in the geometrical configuration of the bundles of collagen under the influence of the elastic fibers may provide a better insight into the mechanisms of distortion of the baroreceptors related to and/or in contact with collagen fibers.     

The distribution and ultrastructure of sensory elements in the baroreceptor region of the truncus arteriosus of the lizard Trachydosaums rugosus

Summary The proximal truncus arteriosus of the lizard Trachydosaurus rugosus was studied with light-, fluorescence and electron-microscopical techniques. Three vessels comprised the truncus: the pulmonary, left aortic, and caroticoaortic arteries. Right and left truncal nerves, each derived from the ipsilateral vagus nerve, innervated the truncus, particularly its proximal 3 mm.Ultrastructurally, the nerves had a variety of appearances: some were clearly adrenergic, c-type or p-type. A number of profiles contained large numbers of mitochondria and were classified as sensory. Some profiles defied exact classification, having characteristics common to two different types of profile.Within the outer medial layers, profiles up to 7 m in diameter were found. These contained large numbers of mitochondria, myelin bodies and structures intermediate between the two. In addition, the profiles contained large amounts of glycogen and small numbers of vesicles. These nerve fibres were classified as baroreceptors, since they closely resemble carotid sinus and aortic arch baroreceptors in mammals.Large numbers of chromaffin cells were found, particularly in the common wall of the pulmonary and left aortic arteries. Many of these cells emitted a long tapering process, which sometimes entered a nearby nerve bundle. Sensory, p-type and c-type profiles, but not adrenergic profiles, made extensive close contacts with chromaffin cells.     

Innervation of the carotid body. An experimental quantitative study

The innervation of the carotid body in the cat was studied by means of light- and electron-microscopic techniques. Sinus nerve resection, glossopharyngeal resection, bilateral cervical sympathectomy, excisions of two nerves, and injection of 6-hydroxydopamine (6-OH-DA) were performed in different groups of animals. It was found that resection of the sinus nerve produces a rapid phase of degeneration of intralobular fibers and synaptic boutons, followed by a reinnervation with a progressive reappearance of these elements. This reinnervation is retarded by sympathectomy and prevented by 6-OH-DA. It is therefore concluded that reinnervation is due to collateral regeneration of nearby sympathetic fibers. Resection of the sinus nerve produces an increase in the number of argentaffin cells and dense-cored vesicles in the cytoplasm of principal cells. These findings suggest the existence of efferent synaptic contacts between this nerve and principal cells. Part of the intralobular fibers and synaptic boutons degenerate after bilateral sympathectomy demonstrating that sympathetic axons connect synaptically to the principal cells. Sympathetic fibers reach the carotid body, not only from branches of the cervical plexuses but also from fibers running in the adventitia of the common carotid artery, and via glossopharyngeal and sinus nerves. The vagus nerve contributes a few fibers to the parenchymal lobules of the carotid body.     

Ultrastructure of calcitonin gene-related peptide- and substance P-like immunoreactive nerve fibres in the carotid body and carotid sinus of the guinea pig

Previous studies have demonstrated that substance P- (SP) and calcitonin gene-related peptide-like immunoreactivities (CGRP-LI) coexist in sensory nerve fibres in the guinea-pig carotid body and carotid sinus. In the present study the ultrastructure of these nerve fibres was investigated by means of single- and double-labelling immunocytochemistry. In both, carotid body and carotid sinus immunoreactive fibres were unmyelinated axons of small diameter (0.12-0.56 microns). At the subcellular level, SP- and CGRP-LI were colocalized in intra-axonal dense core vesicles, suggesting corelease and simultaneous action of these two compounds. SP/CGRP-LI nerve fibres within the carotid body were mainly found in the interparenchymal connective tissue, but also occurred in relationship to blood vessels and nests of glomus cells. Neither in the carotid body not in the carotid sinus, SP/CGRP-LI axons corresponded to the large terminals which are generally considered to represent the main chemoreceptor and baroreceptor endings, respectively. Thus, SP/CGRP-LI fibres either belong to the chemo- and baroreceptors of the C-fibre class or constitute a fibre population not directly involved in conduction of baro- and chemoreflexes.     

Ultrastructure of calcitonin gene-related peptide-and substance P-like immunoreactive nerve fibres in the carotid body and carotid sinus of the guinea pig

Summary Previous studies have demonstrated that substance P-(SP) and calcitonin gene-related peptide-like immunoreactivities (CGRP-LI) coexist in sensory nerve fibres in the guinea-pig carotid body and carotid sinus. In the present study the ultrastructure of these nerve fibres was investigated by means of single-and double-labelling immunocytochemistry. In both, carotid body and carotid sinus immunoreactive fibres were unmyelinated axons of small dianeter (0.12–0.56 m). At the subcellular level, SP-and CGRP-LI were colocalized in intra-axonal dense core vesicles, suggesting corelease and simultaneous action of these two compounds. SP/CGRP-LI nerve fibres within the carotid body were mainly found in the interparenchymal connective tissue, but also occurred in relationship to blood vesslesl and nests of glomus cells. Neither in the carotid body not in the carotid sinus, SP/CGRP-LI axons corresponded to the large terminals which are generally considered to represent the main chemoreceptor and baroreceptor endings, respectively. Thus, SP/CGRP-LI fibres either belong to the chemo-and baroreceptors of the C-fibre class or constitute a fibre population not directly involved in conduction of baro-and chemoreflexes.This study was supported by the DFG, grant He 919/6-2     

The ratio of systemic and coronary fractions of left ventricular output in reflex pressor reactions

In acute experiments the participation of coronary and systemic fractions was studied during suppression of the carotid sinus baroreceptors by the occlusion of the carotid arteries and stimulation of the tibial nerve afferent fibers. In most tests systemic fraction was reduced in carotid arteries occlusion and increased in tibial nerve stimulation. The coronary fraction was always increased. The cardiac output (the sum of systemic and coronary fractions) was steady with blood pressure increase by 20 to 70%. The role of coronary fraction in the mechanism of homeometric regulation of the heart is discussed.     

An electron microscopic study of the baroreceptors in the internal carotid artery of the spontaneously hypertensive rat

Summary The carotid baroreceptor field of normotensive (NTR) and spontaneously hypertensive rats (SHR) examined in this study extends for about 0.5 mm along the length and about 1/3 to 1/2 of the circumference of the wall of the internal carotid artery opposite to the carotid body. The vascular wall of the baroreceptor field exhibits neither a marked dilation to form a carotid sinus nor histological differences in the intima and media compared to other parts of the carotid artery. Histologically the adventitia of the baroreceptor field is characterized by (1) an increased thickness and by less well developed elastic lamellae in comparison with other parts of the arterial wall, (2) a profuse blood and nerve supply, and (3) a richness of cellular elements. The presumptive baroreceptor terminals are localized in the inner 1/3 of the adventitia and display local enlargements that appear to show preferential association with the cell body or processes of the Schwann cell but not with other components of the adventitia. The enlargements are characterized by an accumulation of very densely packed mitochondria, and glycogen particles. No morphological alterations were noted in the baroreceptor terminals of SHR except for proliferated basal laminae that invest the terminals. Our work does not support the concept that resetting of the baroreceptors is due to degeneration of the terminals.Supported by a grant from the Edward G. Schlieder Educational Foundation. Appreciation is extended to Mrs. Lia Pedroza for technical assistance and to Dr. Edward Frohlich of the Alton Ochsner Medical Foundation for supplying the animals used in this research     

Substance P as a baro- and chemoreceptor afferent neurotransmitter: immunocytochemical and neurochemical evidence in the rat

The possibility that substances P (SP) is a neurotransmitter of baro- and chemoreceptor afferents in the rat was investigated. SP-like immunoreactivity (SP-I) was analyzed quantitatively by radioimmunoassay in various levels of the nucleus tractus solitarius (NTS), the site of termination of these afferents while SP-containing afferent neurons were studied in various portions of the peripheral pathways by immunocytochemistry. It was found that the NTS contained significant amounts of SP-I and that unilateral removal of the nodose ganglia reduces the SP-I content of those portions of the NTS known to receive vagal afferents. In addition, SP-I was visualized in discrete fibers in the tunica adventitia of the aortic arch and carotid sinus regions, the vagus nerve and nodose ganglia. These results in the rat are consistent with our previous studies in the cat and provide further evidence that SP is contained within baro- and chemoreceptor afferent nerves.     

Ultrastructure of carotid baroreceptors in the goat.

The initial segment of the occipital artery of the goat appears modified under the electron microscope; endothelial cells are low cuboidal, the tunica media contains many elastic lamellae, and there are few smooth muscle cells. Free afferent nerve endings are seen in this modified arterial wall. They closely resemble the presumptive baroreceptor endings reported in other mammals in their mitochondrial content and abnormal organelles and are interpreted as solely adapting baroreceptors. Special topographical relations between the endings and elastic or collagen fibres do not occur, but microfilaments are present in the vicinity of these endings. Presumptive adrenergic efferent endings are not found in the region of the baroreceptors. We suggest that this zone of the occipited artery is homologous to the carotid sinus.     

FRS2 alpha 2F/2F mice lack carotid body and exhibit abnormalities of the superior cervical sympathetic ganglion and carotid sinus nerve

The docking protein FRS2α is an important mediator of fibroblast growth factor (FGF)-induced signal transduction, and functions by linking FGF receptors (FGFRs) to a variety of intracellular signaling pathways. We show that the carotid body is absent in FRS2α^2F/2F mice, in which the Shp2-binding sites of FRS2α are disrupted. We also show that the carotid body rudiment is not formed in the wall of the third arch artery in mutant embryos. In wild-type mice, the superior cervical ganglion of the sympathetic trunk connects to the carotid body in the carotid bifurcation region, and extends thick nerve bundles into the carotid body. In FRS2α^2F/2F mice, the superior cervical ganglion was present in the lower cervical region as an elongated feature, but failed to undergo cranio-ventral migration. In addition, few neuronal processes extended from the ganglion into the carotid bifurcation region. The number of carotid sinus nerve fibers that reached the carotid bifurcation region was markedly decreased, and baroreceptor fibers belonging to the glossopharyngeal nerve were absent from the basal part of the internal carotid artery in FRS2α^2F/2F mutant mice. In some of the mutant mice (5 out of 14), baroreceptors and some glomus cells were distributed in the wall of the common carotid artery, onto which the sympathetic ganglion abutted. We propose that the sympathetic ganglion provides glomus cell precursors into the third arch artery derivative in the presence of sensory fibers of the glossopharyngeal nerve.     

ULTRASTRUCTURE OF THE CAROTID BODY

An electron microscope investigation was made of the carotid body in the cat and the rabbit. In thin-walled blood vessels the endothelium was fenestrated. Larger vessels were surrounded by a layer of smooth muscle fibers. Among the numerous blood vessels lay groups of cells of two types covered by basement membranes. Aggregates of Type I cells were invested by Type II cells, though occasionally cytoplasmic extensions were covered by basement membrane only. Type I cells contained many electron-opaque cored vesicles (350 to 1900 A in diameter) resembling those in endocrine secretory cells. Type II cells covered nerve endings terminating on Type I cells and enclosed nerve fibers in much the same manner as Schwann cells. The nerve endings contained numerous microvesicles (~500 A in diameter), mitochondria, glycogen granules, and a few electron-opaque cored vesicles. Junctions between nerve endings and Type I cells were associated with regions of increased density in both intercellular spaces and the adjoining cytoplasm. Cilia of the 9 + 0 fibril pattern were observed in Type I and Type II cells and pericytes. Nonmyelinated nerve fibers, often containing microvesicles, mitochondria, and a few electron-opaque cored vesicles (650 to 1000 A in diameter) were present in Schwann cells, many of which were situated close to blood vessels Ganglion cells near the periphery of the gland, fibrocytes, and segments of unidentified cells were also seen. It was concluded that, according to present concepts of the structure of nerve endings, those endings related to Type I cells could be efferent or afferent.     

Reflex mechanisms for changes in renal nerve activity during positive end-expiratory pressure

This study was designed to investigate the interaction between carotid sinus baroreceptors and cardiopulmonary receptors in the reflex control of renal nerve activity (RNA) during positive end-expiratory pressure (PEEP) in anesthetized dogs. PEEP at two different levels (10 and 20 cmH2O) was applied to the following groups: animals with neuraxis intact (I group, n = 12); vagal and aortic nerve denervated animals with carotid sinus nerves intact (V group, n = 6); carotid sinus denervated animals with vagal and aortic nerves intact (SD group, n = 6); and carotid sinus denervated animals also having severed vagal and aortic nerves (SAV group, n = 12). Mean blood pressure (MBP), central venous pressure, and mean airway pressure were also simultaneously measured. In the I group, no significant alterations in RNA occurred during PEEP at both levels, even when MBP fell significantly. Although the drop in MBP in the SD group was similar to that in the I group, RNA decreased significantly 10 s after intervention at both PEEP levels, followed by a recovery of RNA toward the control level. In contrast, a significant increase in RNA, which continued until the end of PEEP, appeared in the V group immediately after each intervention. In the SAV group, RNA responses to PEEP, which were observed in the other groups, were abolished. These results provide evidence that during PEEP, renal nerve activity is modified by an interaction between carotid sinus baroreceptors and cardiopulmonary receptors; excitatory effects occur via carotid sinus nerves and inhibitory effects occur via vagal afferents.     

EFFECTS OF SINUS NERVE STIMULATION ON CAROTID BODY GLOMUS CELLS

The sinus nerve or sympathetic trunk was stimulated unilaterally in one group of adult cats or Syrian hamsters while in another group the sinus nerve or sympathetic trunk was cut unilaterally and the animals were given reserpine. In a third group, atropine was administered prior to sinus nerve stimulation. All tissues were processed for the detection of primary monoamines. The carotid bodies on the operated sides were compared with those on the unoperated sides of the same animal in order to determine if amine depletion occurred following the experimental procedures. After sinus nerve stimulation alone, the density of the granules in the glomus cells was decreased, but changes were not noted in the granules following sympathetic nerve stimulation. Sinus nerve stimulation after atropine administration resulted in no change in granule density. Sinus nerve transection followed by reserpine treatment resulted in a greater decrease in granule density on the unoperated than on the operated side. Transection of the sympathetic components to the carotid body followed by reserpine injections resulted in a decrease in granule density in the glomus cells on both the operated and unoperated sides. These results suggest that the sinus nerve must be intact for reserpine to exert an effect and that the sinus nerve may contain efferent fibers which modulate amine secretion.     

Modulation of baroreceptor activity by gene transfer of nitric oxide synthase to carotid sinus adventitia

Administration of nitric oxide (NO) or NO donors to isolated carotid sinus and carotid bodies inhibits the activity of baroreceptor and chemoreceptor afferent nerves. Furthermore, NO synthase (NOS) is present in endothelial cells and in sensory nerves innervating the carotid sinus region. The major goal of this study was to determine whether overexpression of NOS in carotid sinus modulates baroreceptor activity. Rabbits were anesthetized, and adenoviral vectors (5 x 10(8) plaque-forming units) encoding genes for either beta-galactosidase (beta-Gal) or endothelial type III NOS (eNOS) were applied topically to the adventitial surface of one carotid sinus. In some experiments, the NOS inhibitor N(G)-nitro-l-arginine methyl ester (l-NAME) was applied to the carotid sinus immediately after the vector. Four to five days later, baroreceptor activity and carotid sinus diameter were measured from the vascularly isolated carotid sinus of the anesthetized rabbits. Transgene expression was confirmed by X-Gal staining of beta-Gal and measurement of NOS activity by citrulline assay. The expression was restricted to the carotid sinus adventitia. Baroreceptor activity was decreased significantly, and the pressure-activity curve was shifted to higher pressures in eNOS-transduced (n = 5) compared with beta-Gal-transduced (n = 5) carotid sinuses. The pressure corresponding to 50% of maximum activity averaged 55 +/- 6 and 76 +/- 7 mmHg in beta-Gal- and eNOS-transduced carotid sinuses, respectively (P < 0.05). Decreased baroreceptor activity was accompanied by a significant increase in carotid diameter in the eNOS-transduced carotid sinuses (n = 5). l-NAME prevented the inhibition of baroreceptor activity and the increase in carotid diameter in eNOS-transduced carotid sinuses (n = 5). We conclude that adenoviral-mediated gene transfer of eNOS to carotid sinus adventitia causes sustained, NO-dependent inhibition of baroreceptor activity and resetting of the baroreceptor function curve to higher pressures.     

Three dimensional observation of the nerve fibers along the cerebral blood vessels

Three dimensional observation of the nerve fibers along the cerebral blood vessels was investigated by scanning electron microscopy. Electron probe X-ray microanalysis was also performed in the cerebral blood vessels treated with peroxidase-antiperoxidase immunohistochemistry intensified by nickel ammonium sulfate. Nerve fibers (2-8 microns in diameter) formed a plexus on the outer surface of the adventitia. After branching, the nerve fibers penetrated the blood vessel adventitia. Substance P-immunoreactive nerve fibers showed a meshwork pattern in the outer layer of the adventitia, and vasoactive intestinal polypeptide (VIP)-immunoreactive nerve fibers revealed a spiral running pattern in the inner layer of the adventitia. Taken together with previous studies, these findings suggest that substance P nerve fibers in the cerebral arteries may not be related to arterial dilatation or constriction, but VIP nerve fibers may be vasodilative.     

A method of evoking fast baroreceptor responses for the measurement of the baroreflex latency.

A method for measurement of latency in the baroreflex is described. Standardised square pressure pulse was applied to the isolated carotid sinus and simultaneous recordings of the carotid sinus nerve or sympathetic fibers were performed. Relatively short time of the increase of the pressure makes possible more exact measurements of latency in the baroreflex than other known methods used for this purpose.     

Fine structure of baroreceptor terminals in the carotid sinus of guinea pigs and mice

Summary A light and electron microscopic study was undertaken on the baroreceptor axon terminals in the carotid sinus of guinea pigs and mice, using serial semithin and thin sections.Together with their enveloping Schwann cells, numerous lanceolate axon terminals are organized into a well-defined discoid end organ, referred to as the baroreceptor unit. Baroreceptor units measure 100 to 150 m in diameter and are arranged in a hexagonal pattern. These end organs represent free branched lanceolate mechanoreceptors of complex type (Andres and von Düring, 1973) which belong to the main group of stretch receptors.In the guinea pig the lanceolate terminals enter the media and approach the innermost layers near the intima. In the mouse the terminals are seen to spread in the adventitia and along the medio-adventitial border. Only a few of them penetrate the external elastic layer. Species differences concerning the localization and extent of these visceral mechanoreceptors are discussed, as well as the modified architecture of the sinus wall in the receptor area (elastic segment).Lanceolate terminals form beaded varicosities which are equipped with finger-like or lamellar axoplasmic protrusions. These projections contain a well-differentiated receptor matrix. They are attached to collagen and elastic fibers. The varicosities include densely packed mitochondria, neurotubules, profiles of axoplasmic reticulum, clear and granular vesicles, and striking accumulations of glycogen particles, lamellated bodies and lysosomes. Four types of varicosities are discerned according to their main axoplasmic components. Various types of these varicosities occur within an individual lanceolate terminal.The adrenergic innervation of the carotid sinus was studied by fluorescence histochemistry. In guinea pigs a multilayered wide-meshed plexus of fluorescent fibers occurs in the adventitia where it is closely related to baroreceptor stem fibers. However, adrenergic axons do not enter the media. In mice fluorescent fibers are extremely rare in the adventitia of the carotid sinus.Dedicated to Prof. Dr. Drs.h.c. W. Bargmann on the occasion of his 70th birthdaySupported by a grant from the Deutsche Forschungsgemeinschaft Nr. Bo/525-1. These results were presented in part at the 17. Tagung der Deutschen Gesellschaft für Elektronenmikroskopie, Berlin, Sept. 21.–26., 1975     

Substance P-like immunoreactivity in rat and cat carotid bodies: light and electron microscopic studies

Substance P-immunoreactive (SP-1) structures in the carotid bodies of rats and cats were examined with the light and electron microscopes. In both species SP-I varicose nerve fibers were located singly in the interstitial connective tissue in close association with blood vessels. They were small unmyelinated fibers enveloped in a common Schwann cell sheath with other SP-negative fibers. Some of SP-I fibers contained large dense-cored granules and small clear vesicles in addition to microtubules and mitochondria and probably represented nerve fiber varicosities. The latter often were found incompletely invested by Schwann cell sheaths. SP-fibers were found occasionally in the envelopes of supporting cells at the periphery of parenchymal cell groups. However, none of the nerve terminals making synaptic contacts with glomus cells exhibited SP-like immunoreactivity. In cat carotid bodies some glomus cells showed moderate to intense SP-like immunoreactivity. The intense SP-I glomus cells displayed numerous dense-cored vesicles of 85 to 140 nm in diameter and frequently showed synaptic contacts with SP-negative nerve terminals. In rat carotid bodies we were unable to detect consistent SP-immunoreactivity in glomus cells. Our results do not favor the hypothesis that SP is a neurotransmitter/modulator in the chemoreceptor afferents synapsing on glomus cells in either the cat or rat carotid body. However our results support the hypothesis that SP in cat glomus cells may play a role in the modulation of chemoreceptor activity.     

Ultrastructural studies of the myenteric plexus and smooth muscle in organotypic cultures of the guinea-pig small intestine

External muscle and myenteric plexus from the small intestine of adult guinea-pigs were maintained in vitro for 3 or 6 days. Myenteric neurons and smooth muscle cells from such organotypic cultures were examined at the electron-microscopic level. An intact basal lamina was found around the myenteric ganglia and internodal strands. Neuronal membranes, nuclei and subcellular organelles appeared to be well preserved in cultured tissues and ribosomes were abundant. Dogiel type-II neurons were distinguishable by their elongated electron-dense mitochondria, numerous lysosomes and high densities of ribosomes. Vesiculated nerve profiles contained combinations of differently shaped vesicles. Synaptic membrane specializations were found between vesiculated nerve profiles and nerve processes and cell bodies. The majority of nerve fibres were well preserved in the myenteric ganglia, in internodal strands and in bundles running between circular muscle cells. No detectable changes were found in the ultrastructure of the somata and processes of glial cells. Longitudinal and circular muscle cells from cultured tissue had clearly defined membranes with some close associations with neighbouring muscle cells. Caveolae occurred in rows that ran parallel to the long axis of the muscle cells. These results indicate that the ultrastructural features of enteric neurons and smooth muscle of the guinea-pig small intestine are well preserved in organotypic culture.     

Parasympathetic control of coronary blood flow

Active parasympathetic coronary vasodilation in excess of any changes in myocardial metabolism has been observed in a number of circumstances. Electrical stimulation of the cardiac end of the cut vagus nerve produces a cholinergic coronary vasodilation that is blocked by atropine. Activation of carotid body chemoreceptors, carotid sinus baroreceptors, or left ventricular receptors elicits reflex parasympathetic coronary vasodilation. The coronary vasodilation produced by these reflexes can be prevented by vagotomy or atropine. The relative importance of parasympathetic coronary control in relation to sympathetic and local metabolic coronary control awaits further research.